Mammalian hematopoietic cells provide a diverse range of physiologic activities. These cells are divided into lymphoid, myeloid and erythroid lineages. The lymphoid lineage, comprising B cells and T cells, provides for the production of antibodies, regulation of the cellular immune system, detection of foreign agents in the blood, detection of cells foreign to the host, and the like. The myeloid lineage, which includes monocytes, granulocytes, megakaryocytes, as well as other cells, monitors for the presence of foreign bodies, provides protection against neoplastic cells, scavenges foreign materials, and produces platelets, and the like. The erythroid lineage provides the red blood cells, which act as oxygen carriers.
All publications cited herein are hereby incorporated herein by reference in their entirety.
Despite the diversity of the nature, morphology, characteristics and function of hematopoietic cells, it is presently believed that these cells are derived from a single cell population, termed "stem cells." Unlike more "mature" blood cells, stem cells are capable of self-regeneration but may also divide into progenitor cells that are no longer pluripotent and capable of self-regeneration. These progenitor cells divide repeatedly to form more mature cells which eventually become terminally differentiated to form the various mature hematopoietic cells. Thus the large number of mature hematopoietic cells is derived from a small reservoir of stem cells by a process of proliferation and differentiation. As used herein, "stem cells" refers to hematopoietic stem cells and not stem cells of other cell types.
Stem cells mature into progenitor cells and then become lineage committed, that is, are incapable of maturing into all of the lineages. The use of the words progenitor or progenitor cells indicates cell populations which are no longer stem cells but which have not yet become terminally differentiated. The use of the word lymphoid, myeloid or erythroid in conjunction with progenitor indicates the potential cell populations into which the progenitor is capable of maturing.
Highly purified populations of stem cells currently find use in long-term repopulation of the entire hematopoietic system. Purified progenitor cells of individual lineages would find use only in transiently repopulating or augmenting the various lineages. As progenitors are not believed to be self-regenerating, the repopulation or augmentation would be limited, for example, to short-term hematopoietic reconstitution.
A highly purified or enriched population of stem cells is necessary for a variety of in vitro experiments and in vivo indications. For instance, a purified population of stem cells will allow for identification of growth factors associated with their self-regeneration. In addition, there may be as yet undiscovered growth factors associated with: (1) the early steps of dedication of the stem cell to a particular lineage; (2) the prevention of such dedication; and (3) the negative control of stem cell proliferation.
Stem cells find use in: (1) regenerating the hematopoietic system of a host deficient in any class of hematopoietic cells; (2) a host that is diseased and can be treated by removal of bone marrow, isolation of stem cells and treatment with drugs or irradiation prior to re-engraftment of stem cells; (3) producing various hematopoietic cells; (4) detecting and evaluating growth factors relevant to stem cell self-regeneration; and (5) the development of hematopoietic cell lineages and assaying for factors associated with hematopoietic development.
Stem cells are also important targets for gene therapy, where expression of the inserted genes promotes the health of the individual into whom the stem cells are transplanted. In addition, the ability to isolate stem cells may serve in the treatment of lymphomas and leukemias, as well as other neoplastic conditions where the stem cells are purified from tumor cells in the bone marrow or peripheral blood, and reinfused into a patient after myelosuppressive or myeloablative chemotherapy. Thus, there have been world-wide efforts toward isolating stem cells in substantially pure or pure form.
Stem cells and progenitor cells constitute only a small percentage of the total number of hematopoietic cells. Hematopoietic cells are identifiable by the presence of a variety of cell surface protein or carbohydrate "markers." Such markers may be either specific to a particular lineage or be present on more than one cell type. The markers also change with stages of differentiation. Currently, it is not known how many of the markers associated with differentiated cells are also present on stem and progenitor cells. One marker which was previously indicated as present solely on stem cells, CD34, raised against KG1a cells, is also found on a significant number of lineage committed progenitors. U.S. Pat. No. 4,714,680 describes a composition comprising human CD34.sup.+ stem and progenitor cells.
The CD34 marker is found on numerous lineage committed hematopoietic cells. In particular, 80-90% of the CD34.sup.+ population is marked by other lineage specific and non-specific markers. In view of the small proportion of the total number of cells in the bone marrow or peripheral blood which are stem cells, the uncertainty of the markers associated with the stem cell as distinct from more differentiated cells, and the general difficulty in assaying for human stem cells biologically, the identification and purification of stem cells has been elusive. Characterizations and isolation of human stem cells are reported in: Baum et al. (1992) Proc. Natl. Acad. Sci. USA 89:2804-2808; and Tsukamoto et al. U.S. Pat. No. 5,061,620.
Recently, the mouse stem cell has been obtained in at least highly concentrated, if not a purified form, where fewer than about 30 cells obtained from bone marrow were able to reconstitute all of the lineages of the hematopoietic system of a lethally irradiated mouse. Each assayed cell is multipotent for all hematopoietic lineages, while self-renewal is variable amongst these cells. Spangrude et al. (1988) Science 241:58-62; Smith et al. (1991) Proc. Natl. Acad. Sci. USA 88:2788-2792; Uchida (1992) Ph.D. Thesis Stanford U.; and see also, EPA 89 304651.6 and the references cited therein which describe the isolation of mouse stem cells.
The cell surface marker CDw109, a monomeric GPI-linked glycoprotein of 170 kD has been found to be expressed on primitive T-lymphoblastic leukemias, activated platelets and activated T-lymphoblasts while they are maintained in IL-2. Sutherland et al. (1991) Blood 77:84-93. CDw109 was reported to be unexpressed on whole bone marrow or peripheral blood lymphocytes.